Identification and characterization of mycoviruses capable of inducing hypovirulence in isolates of Sclerotinia sclerotiorum and Rosellinia necatrix

Abstract

newline ix newlineABSTRACT newlineAmongst all plant pathogens fungi cause catastrophic diseases in all major crops with considerable impact on human lives. Sclerotinia sclerotiorum is a cosmopolitan, economically important, and necrotrophic fungal pathogen that infects over majority of the plant species. Rosellinia necatrix, is another diverse fungus that is widely distributed on five continents which is responsible to cause damage to enormous fruit trees and plants. Currently, application of chemical fungicides is the major method used for controlling diseases of important crops. To reduce the dependence on fungicides, highly efficient and environmentally friendly alternative methods are desirable. Therefore, utilizing a biological control method is a tactful choice. The study aimed to isolate and identify mycoviruses which may have the capacity to induce hypovirulence in isolates of Sclerotinia sclerotiorum and Rosellinia necatrix. These mycoviruses in the form of their hypovirulent strains can have the potential to be used as effective biocontrol agents against these pathogens. Sclerotia from infected samples were brought the lab. Sclerotinia spp. were isolated and purified. Similarly, mulberry twigs used as baits, were implanted in infected soil of apple orchards. From baits, Rosellinia was isolated and purified. Isolates showing slow growth phenotype from both the fungal species were considered to be putative hypovirulent and were suspected to be infected with mycovirus. Such isolates were selected and characterized by ITS amplification followed by DNA sequencing, which confirmed the identity of two fungal isolates as Sclerotinia sclerotiorum and Rosellinia necatrix. Presence of a ssDNA mycovirus band from Sclerotinia and a dsRNA mycovirus band from Rosellinia was observed. The nature if these mycoviruses was confirmed as result of different enzymatic treatment viz. DNase, RNase and S1 Nuclease. Further, in order to confirm that the altered phenotype was because of the presence of some mycoviral element; the curing for virus using cycloheximide treatment was done. After curing, cultures exhibited normal morphology, normal growth rate. Also, the mycoviral band seen in the uncured strains was now absent in the cured strains This was followed by the molecular characterization of these isolated mycoviruses. But both these mycoviruses could not be successfully characterized. newlineKeywords: Sclerotinia, Rosellinia, mycovirus, ssDNA virus, dsRNA virus, virus curing.